Presentation on theme: "Colchicine antibodies - Principles of treatment and future"— Presentation transcript:
1 Colchicine antibodies - Principles of treatment and future B. Mégarbane, F.J. BaudRéanimation Médicale et Toxicologique,INSERM U705, Université Paris-Diderot,Hôpital Lariboisière, Paris, France
2 Introduction (1) History: Colchicine is an alkaloid derived from Colchicum autumnale (Liliaceae family)- Native to temperate areas of Europe, Asia, and America- All poisonous plant, high concentrations in the corm or bulbHistory:- Ephemeron (Theophrastlus of Eresus, 370 BC )- Destructive fire of Colchican Medea (Nicander, 150 BC)- Treatment of gout (Alexander of Tralles, 550 AD)- Chemical synthesis (Laborde & Houdé, 1884)
4 Acute colchicine poisoning • Rare in Western countries• Resulting from both accidental and intentional overdoses• Responsible of a high-rate mortality51 None31 Minor19 Moderate3 Major4 Deaths323 case mentions208 single exposuresAAPCC-TESS, Clin Tox 2006355 exposures in 33 countries in Europe during with 4% fatal rateKupferschmidt H, EAPCCT 2005Colchicine toxicity is predictable and dose-dependent with multiorgan involvement and delayed onset.
5 Mechanisms of colchicine action • Selective and reversible binding to the 65 subunits of microtubules- In vitro : high affinity for dividing cells, mitotic apparatus,cilia, sperm tails and brain- Forensic studies: cell blockage in metaphaseand abnormal nuclear morphology• Alteration in multiple cellular functions:- Cell shape, mobility, ability to phagocytosis- Cell division• Affection of organs exhibiting the highest cell turnover(GI mucosa, bone marrow)• Other effects: - collagenase activity- production of PG-B and PG-F2 in synovial cells- transport of nucleosides
6 Toxicokinetics of colchicine Conflicting data ...• A narrow therapeutic index• Rapid absorption with variable bioavailability ( %)• Hepatic metabolism (deacetylation), biliary secretion, enterohepatic cycle• Renal elimination (20%, unchanged form)• Role of co-ingested drugs (CYP3A4 inhibitors, macrolides)Bi-exponential curveElimination half-life : hrsDistribution volume : 21 l/kgRochdi M. Hum Exp Toxicol 1992
7 Colchicine poisoningSigns are well documented, follow a typical pattern, and involve multiorgan systemsStage I (0-12 h) GI symptomsVolume depletionPeripheral leukocytosisStage II (2-7 d) Respiratory distress, ARDS, hypoxemiaCardiovascular shockThromboyctopenia, DICMyelosuppression, neutropeniaHyponatremia, hypocalcemia, hypophosphatemiaMetabolic acidosisRhabdomyolysis, myoglobinuria, oliguric renal failureStage III (1-2 wk) Rebound leukocytosisAlopecia.
8 Experimental assessment of cardiotoxicity Impairment of the intrinsec contractility of rat left ventricular papillary muscle(2 and 4mg/kg colchicine versus saline)• Impairment of myocardial contractility: maximum shortening velocity (-32% and -61%) active isometric force (-47% and -65%) peak power output (-57% and -69%)• Impairment of isotonic relaxation and load dependence of relaxation, suggesting a decrease in sarcoplasmic reticulum function• Acceleration of isometric relaxation, suggesting a decrease in Ca++ myofilament sensitivity• Marked negative inotropic effects if impairment of myothermal economyMery P. Intensive Care Med 1994
9 Colchicine cardiac toxicity Cardiovascular and direct negative inotropic effects were largely assessed.2 series with very elevated mortality rate:• Bismuth C. Presse Med 1977:11 cases ≥ 0.8mg/kg GI troubles + DIC + aplasia + cardiac shock = 100% death• Sauder P. Hum Toxicol 1983:8 cases mg, 4 cases with cardiac failure ( cardiac index, systemic resistance) = 100% deathFew cases report with survive despite cardiogenic shock:• Baron DA. Ouest Med F 38 yrs, 30 mg, WBC 27,000 /mm3, FV 22%• Bismuth C. Nouv Presse Méd M 37 yrs, 0.8 mg/kg, WBC 12,000 /mm3, PT 19%• De Villota E. Crit Care Med M 50 yrs, 30 mg, WBC 9,100 /mm3
10 Assessment of colchicine mechanism of cardiac toxicity Sauder P. Hum Toxicol, 1983
11 T-waves negativation, ST elevation in leads I, II V2-V6 Profound EKG changes :Colchicine may impair impulse generation and cardiac conduction, in addition to the electrolyte and acid-base disturbancesCardiac dysrhythmias(sinus tachycardia, sinus bradycardia, VF, sinus arrest, complete AV blockade)T-waves negativation, ST elevation in leads I, II V2-V6Stahl. Am J Med Sci 1979Stapczynski. Ann Emer Med 1982Hobson. Anaesth Intensive Care 1986Wells. Vet Hum Toxicol 1989Stemmermann. Hum Pathol 1972McIntyre. J Forensic Sci 1994Weakley-Jones. Am J Forensic Med Pathol 2001Wells. Vet Hum Toxicol 2000Brvar. Wien Klin Wochenschr 2004Murray. Mayo Clin Proc 1983Mendis. Postgraduate J 1989Wells. Vet Hum Toxicol 1989Mullins. Am J Emerg Med 2000Weakleyy-Jones. Am J Forensic Med Pathol 2001Brvar. Crit Care 2003Miller. J Emerg Med 2005Van Heyningen C. Emerg Med J 2005
12 Direct colchicine-related cardiac injuries • Biology alterations: Delayed elevation of myocardial enzymes (troponine, CPK-MB)Mullins ME. Am J Emerg Med 2000Sussman JS. Ther Drug Monit 2004Van Heyningen C. Emerg Med J 2005• Pathology: a series of 12 fatal casesInterstitial edema without cell necrosis in all cases.Interstitial myocarditis in 2 cases.Hoang C. Ann Pathol 1982
13 Prognostic factors • The supposed ingested dose Mortality < 0.5 mg/kg < 5 %0.5 à 0.8 mg/kg %> 0.8 mg/kg 90 %• Prothrombin index ≤ 20%• WBC ≥ 18x109 /l• Onset of cardiogenic shock• Onset of ARDSin the 24th hoursin the 72th hoursBismuth C. Presse Med 1977
14 Colchicine poisoning management • Management includes early GI decontamination, careful monitoring of physical examination and laboratory tests (electrolytes, blood gases, appropriate cultures) and supportive treatments:• To date, there is no successful commercially available specific therapy, although several experimental studies and one human case report assessed the efficiency of colchicine-specific Fab fragments.Fluids (diarrhea)Vasopressors (shock)Oxygen supplementation, mechanical ventilation (ALI, ARDS)Antibiotics (fever)GCSF (neutropenia)Transfusions (thrombocytopenia, anemia)
15 Principles of immunotoxicotherapy • A procedure able to simultaneously sequester, extract or redistribute, and eliminate the toxin by using specific active binding sites derived from different antibody molecular entities.• Currently used in humans for treating cardiac glycoside and venom poisonings. Since the initial report in 1976 by Smith et al., it has become the first-line treatment of life-threatening intoxication by cardiac glycosides, including digoxin, digitoxin, and other structurally related cardiotoxins from Nerium, Thevetia sp. (oleander), and Bufo sp. (toads).Scherrmann JM. Clin Toxicol 1989
16 Specific active biding sites derived from antibody • Industrial manufacturing• Polyclonal antibodies of ovine origin• Fragmentation of antibodies:- useful for haptens (< 1000 d)- increased safety- limited instability• Future trends: monoclonal, humanized, semi-synthetic, further fragmentation, ...
17 Toxin sequestration Pharmacokinetic characteristics: Affinity issues: • Vdtoxin/Vdantitoxin ratio 1 to probability of interactionFor toxins with vascular distribution: IgG, IgM (5 l)For toxin with extravascular distribution: Fab (30 l)• Equilibration time in the distribution space: 2-4h (Fab) versus h (IgG, Fab’2)Affinity issues:• Affinity ≥ 109 M-1 (critical minimal value) to form stable complexes• Prefer polyclonal to monoclonal antibodies, based on manufacturing capacity + to enlarge specificity to epitopes.• Prefer ovine to equine antibodies, to reduce serum sickness + to produce a higher proportion of specific IgG
18 Toxin extraction or redistribution Involve the removal of toxin from the toxin receptor compartment to the antibody distribution spaceLimiting factors:- Reversibility of binding (reversible intracellular binding)- Kinetics of the toxin release from the receptort1/2 dissociation of colchicine / tubulin : 20 ht1/2 dissociation of digoxin / Na,K-ATPase: 1 hBlood anti-toxin concentration should exceed bound toxins during a predicted period of time to allow toxin effluxAnti-toxin with slow clearance (IgG, Fab ’2 < Fab) should be preferred to allow redistribution over a prolonged period
19 Toxin eliminationLow MW-toxin bound to anti-toxin adopt anti-toxin elimination propertiesExtra-renal clearance: = 60-70%Reticulo-endothelial tissuesgut, liver, spleen, lymph nodesRenal clearance:• Fab are filtrated through glomeruli, rapidly and extensively reabsorbed and catabolized by the proximal tubule cells.• Fab increases the renal clearance of toxins: %However: Possible tubular re-absorptionRe-circulation of free toxin moleculesReduction in glomerular filtration
21 Developmental requisites for immunotoxicotherapy Colchicine is a good candidate for the development of a successful immunotherapy, as it perfectly answers to the toxin-dependent requisites.1- Severe poisonings with a high risk of death and short-term effects2- Toxicity in the milligram range, allowing stoechiometric neutralization3- Efficient production of antibody in animals after conjugation to a protein4- High affinity antibody ( M-1)5- Distribution volume much greater than its corresponding Fab with the possibility of a rapid redistribution from tissue to blood
22 Limitations in colchicine poisoning (1) Colchicine characteristics may theoretically limit the potential benefitsWhereas cardiac glycosides are membrane-associated toxins, efficiently reversed with an equimolar dose of specific Fab colchicine is low-MW acting intracellular poison, needing to consider an expanding immunotherapy model to intracellular toxins.
23 Limitations in colchicine poisoning (2) By opposite to digitalis poisonings where only one organ, the heart, is at vital risk, severe colchicine poisonings induce multi-organ failure with not always reversible structural injuries.• There is no evidence that antibodies can reverse organ injuries.• In order to consider any therapeutic interest of antibodies, it is necessary to hypothesize that neutralization of residual unbound colchicine may convert a lethal intoxication into a disease compatible with survival, even if a greater part of the toxin has already damaged the organism.
24 Assessment of the efficacy of toxin detoxification using immunotoxicotherapy To test the reversal of toxicityTo test the sequestration effectTo test the best fragments and administration conditionsScherrmann JM. Clin Toxicol 1989
25 In vitro experimental studies (1) Partial reversion of colchicine-induced inhibition of tubulin polymersiation using colchicine-sepcific antibodiesWolf AD. J Biol Chem 1980
26 In vitro experimental studies (2) Colchicine neutralization with antibodies has proven to be effective. Although intracellular binding of colchicine to microtubules was expected to limit immunotoxicotherapy efficacy, a reversible effect on microtubules was found.• Colchicine-induced polyploidy and chromosomal aberrations in a model of Chinese hamster ovary cell were reversible with a specific high-affinity monoclonal antibody, even when administered up to 6 hours after colchicine exposureRouan SE. Am J Pathol 1990 • A tightly bound intracellular toxin was extracted with high-affinity antibodies at a rate depending on its dissociation rate from its receptors. Kinetics was of first-order decline with t1/2 ranging from 15.5 to 16.4 hChappey ON. J Pharmacol Exp Ther 1995
27 In vivo experimental studies (1) Anti-colchicine active immunization of rabbits:- Protective effects against 3-mg/kg colchicine (> LD50), with an antibody titer-dependent response.- Effective trapping of colchicineBiexponential - t1/2: 12hUnbound colchicine < 0.5%Scherrmann JM. Clin Toxicology 1989
28 In vivo experimental studies (2) In mice, colchicine-specific IgG administration, even after colchicine distribution phase, significantly decreased mortality rate.Terrien N. Toxicol Appl Pharmacol 1990In mice, colchicine-specific goat IgG (1/2-1/8 molar dose) favorably improved outcome when previously (90 min) receiving IP 3.8-mg/kg colchicine lethal dose.Sabouraud AE. Toxicology 1991
29 Colchicne Fab fragments-related alteration in kinetics Tissue extraction & elimination:In mice: Fab induced• Decrease in colchicine VD• Colchicine sequestration in intravascular spaces• Decrease of colchicine concentrations in most tissuesSabouraud AE. J Pharm Pharmacol 1992In rabbits: Fab infusion 1.5 h after 0.1-mg/kg colchicine, over 0.25 h, 1/2-molar doseColchicine concentration x10-16 within 15 minTotal plasma AUC x20Undetectable free plasma fraction over a period of 2 h• Decrease of the VD /24• Decrease of total clearance: /17• Reduction of biliary excretion: - 80%• Reduction of the dose fraction excreted in urine: 9% versus 38%Sabouraud AE. J Pharmacol Exp Ther 1992
30 Case report of a severe human colchicine overdose treated with colchicine-specific Fab fragments (1) 25-year-old woman, suicidal attempt, 0.96 mg/kg (60 mg) colchicine.- Call: 24 h after ingestion due to severe GI pain- On the scene: HR 110 /min, unrecordable BP500 ml colloid and 10 mg/kg/min dobutamine- ICU: °C, BP 110/80 mmHg, HR 110 /min, RR 60 /min,Creatinine 140 µmol/l,WBC 69,300 /mm3, platelet 268,000 /mm3, PT 14%, V <5%pH 7.38, PaCO2 28 mmHg, PaO2 86 mmHg (O2 3 l/min)Chest X-Rays: pulmonary edemaPlasma colchicine: 24 ng/ml (RIA)Gastric lavage + activated charcoalCrystalloid (4400 ml) and colloid (1100 ml)Deterioration of hemodynamic status despite dobutamineBaud FJ. N Engl J Med 1995
31 Case report (2) -45 min -15 min +1H +12H PAP (mmHg) 23 18 13 11 PCWP (mmHg)SBP (mmHg)HR (/min)Cardiac Index (l/min/m2)Resistance (dyn.sec.cm-5/m2)AV difference in OPlasma lactate (mmol/l) NDDobutamine (µg/kg/min)Dopamine (µg/kg/min)Norepinephrine (mg/h)Baud FJ. N Engl J Med 1995
32 Infusion of goat colchicine-specific Fab fragments H36 after ingestion Case report (3)Infusion of goat colchicine-specific Fab fragments H36 after ingestion• Antiserum derived from goats immunized with a conjugate of colchicine + serumalbumine• Affinity: 2 x 1010 M-1• No separation of colchicine-specific Fab fragments (7.5%) from others• Dosage: 480 mg (6.4 g Fab fragments) in 160 ml240 mg over 1-hour period240 mg over 6 hoursBaud FJ. N Engl J Med 1995
33 Case report (4) -45 min -15 min +1H +12H PAP (mmHg) 23 18 13 11 PCWP (mmHg)SBP (mmHg)HR (/min)Cardiac Index (l/min/m2)Resistance (dyn.sec.cm-5/m2)AV difference in OPlasma lactate (mmol/l) NDDobutamine (µg/kg/min)Dopamine (µg/kg/min)Norepinephrine (mg/h)Baud FJ. N Engl J Med 1995
34 Case report (5) Toxicodynamics: • Significant arterial vasodilatation within 30 min.• Marked improvement in all indexes of tissue perfusion.• Withdrawal of inotropic drugs within 36 h.• ARDS took longer to subside.• No prevention of the delayed occurrence of bone marrow aplasia, complete hair loss, and transient peripheral neuropathy.• No adverse effect (hypersensitivity, serum sickness) or recurrence of toxic signsSurvival with the well-known sequential phases however with a shortened durationBaud FJ. N Engl J Med 1995
35 Case report (6) Toxicokinetics: • total plasma concentration (x6: 12 to 122 ng/ml), as soon as 10 min after infusion, whereas free concentration became undetectable• urinary excretion (x6), initially bound to Fab fragmentsRenally-excreted colchicine: 5.2 mg• Neutralized colchicine = 3.7 mg / 9 mg of present colchicine• Fab/total colchicine ratio: 4 (0-1 h), 2 (1-7h) and <1 (>7h)T1/2: 25 hBaud FJ. N Engl J Med 1995
36 Case report (7)The patient’s clinical improvement was explained by the direct effects of colchicine-specific Fab fragments:• Redistribution from tissues into plasmaIncrease in plasma concentration• Sequestration into plasma compartmentHigh affinity of Fab to colchicine• Immunoneutralization of the effectsDecrease of protein-unbound colchicine (during 7h-infusion)and subsequent partial rebound (H12)Baud FJ. N Engl J Med 1995
37 The situation today?To date, no commercial preparation of colchicine-specific Fab fragments is available.Why ?Solved problems:• Specific antibody development is difficult• Fragmentation technique is a complex procedure• Large amount of fragments is required for a stoechiometrical neutralizationUnsolved problems:- Inadequation between development costs and potential prescriptions (orphan)- Difficulties (legislation and bureaucracy) to organize an European trial
38 Further considerations for colchicine immunotoxicotherapy Regarding the indications and time to administer:• Immediate or potentially life-threatening intoxication• Concept of critical dose (when a risk of mortality occurs)• Challenge the classical thinking of the necessity of rapid administration of the total Fab dose to improve poisoning.Regarding the minimal efficient dose to administer:• Optimal amount to neutralize = total ingested dose - critical dose• Inframolar neutralization may be efficient as in animal reports.• Removal of a modest portion of colchicine may dramatically improve outcome.Regarding the therapeutic action:Further studies are needed to determine the Fab effect on bone marrow aplasia.Baud FJ. Arch Toxicol Suppl 1997
39 Conclusions:• Coclchicine-specific Fab fragments may be useful to complete the supportive care in the most severe colchicine poisonings.• However, to date, these fragments are still not commercially available.• Until issues of cost and supply are worked out, coclchicine-specific Fab fragments remain a dream in the areas where poisonings are frequent and specific therapy urgently required.